The long term objective of this project is to obtain an understanding of the mechanisms by which the activities of disparate genes are temporally controlled. The system I have chosen as a model of such regulation is the production of the chorion (eggshell) by the monolayer of follicle cells surrounding the maturing oocytes of the commericial silkmoth, Bombyx mori. In these cells the synthesis of well over 100 chorion proteins follows a precise temporal pattern corresponding to a program of specific mRNA production. The majority of these chorion proteins are members of large multigene families. At least 50 percent of these genes have been localized to continuous segments of DNA 370 kb in length. The remaining chorion genes will be cloned and linked to these segments using modified chromosomal walking procedures. The individual genes encountered in the locus will be classified as to their respective gene family and to their precise period of expression in choriogenesis. Remarkable variability of the DNA level has been detected between different inbred strains of B. mori. A detailed description of this variation will be obtained by additional chromosomal walking in these strains. Any conserved features revealed by this analysis will indicate elements critical to the maintenance of coordinate gene expression. The variable features will suggest the processes by which the gene families can change during evolution. Two possible mechanism for the regulation of chorion gene expression will be tested. Chromatin accessibility will be explored by monitoring developmental changes in DNase sensitivity, state of methylation or association with the nuclear matrix. Regulation by means of promoter structure will be explored by comparative DNA sequence determination of the ca. 250 bp 5' regions separating the divergently oriented chorion gene pairs.